Carbonic anhydrase IX is a pH-stat that sets an acidic tumour extracellular pH in vivo

Background Tumour Carbonic Anhydrase IX (CAIX), a hypoxia-inducible tumour-associated cell surface enzyme, is thought to acidify the tumour microenvironment by hydrating CO2 to form protons and bicarbonate, but there is no definitive evidence for this in solid tumours in vivo. Methods We used 1H magnetic resonance spectroscopic imaging (MRSI) of the extracellular pH probe imidazolyl succinic acid (ISUCA) to measure and spatially map extracellular pH in HCT116 tumours transfected to express CAIX and empty vector controls in SCID mice. We also measured intracellular pH in situ with 31P MRS and measured lactate in freeze-clamped tumours. Results CAIX expressing tumours had 0.15 pH-unit lower median extracellular pH than control tumours (pH 6.71 tumour vs pH 6.86 control, P = 0.01). Importantly, CAIX expression imposed an upper limit for tumour extracellular pH at 6.93. Despite the increased lactate concentration in CAIX-expressing tumours, 31P MRS showed no difference in intracellular pH, suggesting that CAIX acidifies only the tumour extracellular space. Conclusions CAIX acidifies the tumour microenvironment, and also provides an extracellular pH control mechanism. We propose that CAIX thus acts as an extracellular pH-stat, maintaining an acidic tumour extracellular pH that is tolerated by cancer cells and favours invasion and metastasis.We are grateful for the support of CRUK [grant number C14303/A17197], the Breast Cancer Research Foundation, the Royal Society, Worldwide Cancer Research and the European Research Council [SURVIVE: 723397]. JP-T and SC received support from the Spanish Ministry of Economy and Competitiveness SAF2014-23622